Liver X Receptor Agonist T0901317 Research Articles

LXR Agonist T0901317's Hepatic Impact Overrules Its Atheroprotective Action in Macrophages, Driving Early Atherogenesis in Chow-Diet-Fed Male Apolipoprotein E Knockout Mice.

Preclinical studies regarding the potential of liver X receptor (LXR) agonists to inhibit macrophage foam cell formation and the development of atherosclerotic lesions are generally executed in mice fed with Western-type diets enriched in cholesterol and fat. Here, we investigated whether LXR agonism remains anti-atherogenic under dietary conditions with a low basal hepatic lipogenesis rate. Hereto, atherosclerosis-susceptible male apolipoprotein E knockout mice were fed a low-fat diet with or without 10 mg/kg/day LXR agonist T0901317 supplementation for 8 weeks. Importantly, T0901317 significantly stimulated atherosclerosis susceptibility, despite an associated increase in the macrophage gene expression levels of cholesterol efflux transporters ABCA1 and ABCG1. The pro-atherogenic effect of T0901317 coincided with exacerbated hypercholesterolemia, hypertriglyceridemia, and a significant rise in hepatic triglyceride stores and macrophage numbers. Furthermore, T0901317-treated mice exhibited elevated plasma MCP-1 levels and monocytosis. In conclusion, these findings highlight that the pro-atherogenic hepatic effects of LXR agonism are dominant over the anti-atherogenic effects in macrophages in determining the overall atherosclerosis outcome under low-fat diet feeding conditions. A low-fat diet experimental setting, as compared to the commonly used high-fat-diet-based preclinical setup, thus appears more sensitive in uncovering the potential relevance of the off-target liver effects of novel anti-atherogenic therapeutic approaches that target macrophage LXR.

Liver X receptor agonist upregulates LPCAT3 in human aortic endothelial cells.

Endothelial cells (ECs) play an important role in tissue homeostasis. Recently, EC lipid metabolism has emerged as a regulator of EC function. The liver X receptors (LXRs) are involved in the transcriptional regulation of genes involved in lipid metabolism and have been identified as a potential target in cardiovascular disease. We aimed to decipher the role of LXRs in the regulation of lipid metabolism in human aortic endothelial cells. Lipid composition analysis of endothelial cells treated with the LXR agonist T0901317 revealed that LXR activation increased the proportion of polyunsaturated fatty acids (PUFAs) and decreased the proportion of saturated fatty acids. The LXR agonist decreased the uptake of fatty acids (FAs) by ECs. This effect was abolished by LXRα silencing. LXR activation increased the activity and the expression of lysophosphatidylcholine acyltransferase, LPCAT3, which is involved in the turnover of FAs at the sn-2 position of phospholipids. Transcriptomic analysis also revealed that LXRs increased the expression of key genes involved in the synthesis of PUFAs, including FA desaturase one and 2, FA elongase 5 and fatty acid synthase. Subsequently, the LXR agonist increased PUFA synthesis and enhanced arachidonic acid, eicosapentaenoic acid, and docosahexaenoic acid content in the EC phospholipids. Modification of the FA composition of ECs by LXRs led to a decrease of arachidonate and linoleate derived prostaglandins synthesis and release. No change on markers of inflammation induced by plasma from sickle cell patient were observed in presence of LXR agonist. These results identify LXR as a key regulator of lipid metabolism in human aortic endothelial cells and a direct effect of LXR agonist on lysophosphatidylacyl transferase (LPCAT3).

Ginsenoside F2 Restrains Hepatic Steatosis and Inflammation by Altering the Binding Affinity of Liver X Receptor Coregulators

BackgroundGinsenoside F2 (GF2), the protopanaxadiol-type constituent in Panax ginseng, has been reported to attenuate metabolic dysfunction-associated steatotic liver disease (MASLD). However, the mechanism of action is not fully understood. Here, this study investigates the molecular mechanism by which GF2 regulates MASLD progression through liver X receptor (LXR). MethodsTo demonstrate the effect of GF2 on LXR activity, computational modeling of protein-ligand binding, Time-resolved fluorescence resonance energy transfer (TR-FRET) assay for LXR cofactor recruitment, and luciferase reporter assay were performed. LXR agonist T0901317 was used for LXR activation in hepatocytes and macrophages. MASLD was induced by high-fat diet (HFD) feeding with or without GF2 administration in WT and LXRα−/− mice. ResultsComputational modeling showed that GF2 had a high affinity with LXRα. LXRE-luciferase reporter assay with amino acid substitution at the predicted ligand binding site revealed that the S264 residue of LXRα was the crucial interaction site of GF2. TR-FRET assay demonstrated that GF2 suppressed LXRα activity by favoring the binding of corepressors to LXRα while inhibiting the accessibility of coactivators. In vitro, GF2 treatments reduced T0901317-induced fat accumulation and pro-inflammatory cytokine expression in hepatocytes and macrophages, respectively. Consistently, GF2 administration ameliorated hepatic steatohepatitis and improved glucose or insulin tolerance in WT but not in LXRα−/− mice. ConclusionGF2 alters the binding affinities of LXRα coregulators, thereby interrupting hepatic steatosis and inflammation in macrophages. Therefore, we propose that GF2 might be a potential therapeutic agent for the intervention in patients with MASLD.

Administration of an LXR agonist promotes atherosclerotic lesion remodelling in murine inflammatory arthritis.

The leading cause of mortality in patients with rheumatoid arthritis is atherosclerotic cardiovascular disease (CVD). We have shown that murine arthritis impairs atherosclerotic lesion regression, because of cellular cholesterol efflux defects in haematopoietic stem and progenitor cells (HSPCs), causing monocytosis and impaired atherosclerotic regression. Therefore, we hypothesised that improving cholesterol efflux using a Liver X Receptor (LXR) agonist would improve cholesterol efflux and improve atherosclerotic lesion regression in arthritis. Ldlr -/- mice were fed a western-type diet for 14 weeks to initiate atherogenesis, then switched to a chow diet to induce lesion regression and divided into three groups; (1) control, (2) K/BxN serum transfer inflammatory arthritis (K/BxN) or (3) K/BxN arthritis and LXR agonist T0901317 daily for 2 weeks. LXR activation during murine inflammatory arthritis completely restored atherosclerotic lesion regression in arthritic mice, evidenced by reduced lesion size, macrophage abundance and lipid content. Mechanistically, serum from arthritic mice promoted foam cell formation, demonstrated by increased cellular lipid accumulation in macrophages and paralleled by a reduction in mRNA of the cholesterol efflux transporters Abca1, Abcg1 and Apoe. T0901317 reduced lipid loading and increased Abca1 and Abcg1 expression in macrophages exposed to arthritic serum and increased ABCA1 levels in atherosclerotic lesions of arthritic mice. Moreover, arthritic clinical score was also attenuated with T0901317. Taken together, we show that the LXR agonist T0901317 rescues impaired atherosclerotic lesion regression in murine arthritis because of enhanced cholesterol efflux transporter expression and reduced foam cell development in atherosclerotic lesions.

T0901317, a liver X receptor agonist, ameliorates perinatal white matter injury induced by ischemia and hypoxia in neonatal rats

Perinatal white matter injury (PWMI) can lead to permanent neurological damage in preterm infants and bring a huge economic burden to their families and society. Liver X receptors (LXRs) are transcription factors that have been confirmed to mediate the myelination process under physiological conditions and are involved in regulating neurogenesis in adult animal models of acute and chronic cerebral ischemia. However, the role of LXRs in PWMI induced by both ischemic and hypoxic stimulation in the immature brain has not been reported. Herein, we investigated the role of LXRs in a neonatal rat model of white matter loss after hypoxia-ischemia (HI) injury through intraperitoneal injection of the LXR agonist T0901317 (T09) 1 day before and 15 min postinjury. The in vivo data showed that T09 treatment significantly facilitated myelination and ameliorated neurological behavior after PWMI. Moreover, T09 enhanced the proliferation of oligodendrocyte lineage cells and reduced microgliosis and astrogliosis in the microenvironment for oligodendrocytes (OLs), maintaining a healthy microenvironment for myelinating OLs. In vitro data suggested that the expression of the myelin-related genes Plp and Cnpase was increased in OLN-93 cells after T09 intervention compared with OLN-93 cells injured by oxygen and glucose deprivation (OGD). In primary mixed astrocytes/microglia cells, T09 also reduced the expression of Il6, Cox2, Tnfa and Il10 that was induced by OGD. Mechanistically, the mRNA expression level and the protein level of ATP binding cassette subfamily A member 1 (Abca1) decreased after HI injury, and the protective effect of T09 might be related to the activation of the LXRβ-ABCA1 signaling pathway. Our study revealed the protective role of LXRs in myelination and white matter homeostasis, providing a potential therapeutic option for PWMI.

LXR agonist inhibits inflammation through regulating MyD88 mRNA alternative splicing.

Liver X receptors (LXRs) are important regulators of cholesterol metabolism and inflammatory responses. LXR agonists exhibit potently anti-inflammatory effects in macrophages, which make them beneficial to anti-atherogenic therapy. In addition to transrepressive regulation by SUMOylation, LXRs can inhibit inflammation by various mechanisms through affecting multiple targets. In this study, we found that the classic LXR agonist T0901317 mediated numerous genes containing alternative splice sites, including myeloid differentiation factor 88 (MyD88), that contribute to inflammatory inhibition in RAW264.7 macrophages. Furthermore, T0901317 increased level of alternative splice short form of MyD88 mRNA by down-regulating expression of splicing factor SF3A1, leading to nuclear factor κB-mediated inhibition of inflammation. In conclusion, our results suggest for the first time that the LXR agonist T0901317 inhibits lipopolysaccharide-induced inflammation through regulating MyD88 mRNA alternative splicing involved in TLR4 signaling pathway.

Liver X receptor agonists exert antitumor effects against hepatocellular carcinoma via inducing REPS2 expression.

Recent studies show that liver X receptor (LXR) agonists exert significant antitumor effects in a variety of tumor cell lines including hepatocellular carcinoma (HCC). But the molecular mechanisms underlying LXR antitumor activity are not fully understood. In this study we investigated the effect of LXR agonist T0901317 (T317) on HCC development and its relationship with RalA binding protein 1 (RALBP1)-associated EPS domain containing 2 (REPS2)/epidermal growth factor receptor (EGFR) signaling axis. We showed that T317 (0.1-0.5 μM) dose-dependently increased REPS2 expression in normal hepatocytes (BNLCL.2 and LO2) and HCC cells (HepG2 and Huh-7). Using promoter activity assay and chromatin immunoprecipitation (CHIP) assay we demonstrated that T317 enhanced REPS2 expression at the transcriptional level via promoting the binding of LXR protein to the LXR-response element (LXRE) in the REPS2 promoter region. We showed that the inhibitory effect of T317 on the proliferation and migration of HCC cells was closely related to REPS2. Moreover, we revealed that T317 (400 nM) increased expression of REPS2 in HepG2 cells, thus inhibiting epidermal growth factor (EGF)-mediated endocytosis of EGFR as well as the downstream activation of AKT/NF-κB, p38MAPK, and ERK1/2 signaling pathways. Clinical data analysis revealed that REPS2 expression levels were inversely correlated with the development of HCC and reduced REPS2 expression associated with poor prognosis, suggesting that REPS2 might beinvolved in the development of HCC. In conclusion, this study provides new insights into the potential mechanisms of LXR agonist-inhibited HCC.

LXRβ Activation Inhibits the Proliferation of Small-cell Lung Cancer Cells by Depleting Cellular Cholesterol.

Liver X receptors (LXRs) are nuclear receptors with various functions, including the regulation of cholesterol metabolism, glucose homeostasis, and inflammation. We previously reported that LXR activation inhibits the growth of oral cancer cells by inducing cellular cholesterol efflux and that LXRβ is expressed mainly in small-cell lung cancer (SCLC) tissues. SCLC is one of the most aggressive cancers, and identifying an effective therapeutic target molecule is desirable. Therefore, we investigated whether LXRβ could be an effective target molecule for SCLC treatment through in vitro experiments. We evaluated the influence of treatment with the LXR agonist T0901317 on cell proliferation and apoptosis in SCLC cell lines using cell viability, BrdU-ELISA, FACS, and western blot analyses. Moreover, the mechanism by which T0901317 inhibits SCLC cell proliferation was elucidated using qRT-PCR, western blot, a cholesterol quantification assay, and a genome editing technique. We showed that cultivated SCLC cells expressed LXRβ and that an LXR agonist inhibited the proliferation of SCLC cells without toxicity to normal cells. Furthermore, the antitumoral effect of an LXR agonist on SCLC cells was attributed to the induction of ABCA1 by LXRβ activation, resulting in an increase in cellular cholesterol efflux via ABCA1. The activation of LXRβ up-regulates ABCA1 expression, causing cholesterol depletion in cancer cells. This mechanism could be a novel target strategy for SCLC.

Design, synthesis and structure–activity relationship of 4-(1,1,1,3,3,3-hexafluoro-2-hydroxyisoprop-2-yl)phenylsilane derivatives as liver X receptor agonists

Liver X receptor (LXR) α and LXRβ are nuclear receptors playing key roles in lipid metabolism, and LXR ligands are attractive drug candidates for metabolic disorders. Here we report the structural development of 4-(1,1,1,3,3,3-hexafluoro-2-hydroxyprop-2-yl)phenylsilane derivatives as LXR agonists bearing silyl functionalities as the hydrophobic pharmacophore, based on the structure of the known sulfonamide LXR agonist T0901317. Most of the synthesized compounds exhibit agonistic activity toward LXRs, but the LXR subtype-selectivity differs depending upon the substituents on the silicon atom. Among them, tri(n-propyl) derivative 12 shows potent LXR-agonistic activity with moderate α subtype-selectivity, while dimethylphenylsilyl derivative 19 shows modest β-selectivity. These results indicate that silanes can serve as an alternative to the sulfonamide moiety of LXR agonists, and are promising structural options for the development of novel subtype-selective LXR agonists.

The liver X receptor agonist T0901317 reduces the inflammation of alveolar epithelial cells induced by polyhexamethylene guanidine through inhibition of the NFκB signaling pathway

BackgroundAs a kind of disinfectant, polyhexamethylene guanidine (PHMG) can cause pulmonary inflammation. In addition to liver X receptors (LXRs) playing an important role in cholesterol and lipid metabolism, it has also been found to be involved in inflammation in recent years. This article explores the role of LXRs agonist T0901317 in the inflammation of alveolar epithelial cells induced by PHMG.MethodsThe A549 human alveolar basal epithelial cell line was exposed to PHMG, T0901317, or the nuclear factor (NF)κB inhibitor BAY11-7082. The cell survival rate was used to determine the cytotoxicity of PHMG and T0901317 to A549 cells. Western blot analysis was used to determine the expression of proteins related to the LXRs and the NFκB signaling pathway. Enzyme-linked immunosorbent assay (ELISA) was conducted to examine the expression of inflammatory cytokines such as interleukin (IL)-8 and interleukin (IL)-6.ResultsIncubation of A549 cells with PHMG decreased the expression of LXRs-related proteins, reduced the expression of cellular IκB, increased the expression of nuclear NFκB, and increased the levels of the inflammatory cytokineIL-8 and IL-6. However, pretreatment with the LXR agonist T0901317 partially reversed the effects of PHMG. The effects of T0901317on NFκB signaling pathway was similar to that observed with the NFκB inhibitor BAY11-7082.ConclusionsThe LXRs agonist T0901317 may reduce the inflammation of alveolar epithelial cells induced by PHMG by inhibiting the NFκB signaling pathway.

Activation of liver X receptors ameliorates alterations in testicular function in rats exposed to electromagnetic radiation

ABSTRACT Introduction: Cell phones are essential for modern life and their usage becomes prevalent. Worries have increased concerning the harmful effects of their radio-frequency electromagnetic radiation. Liver X receptors (LXRs) are ligand-dependent transcription factors that have important roles in lipid metabolism and inflammation, plus several recently emerging roles. Whether it is useful to overcome the hazardous effects of cell phone radiation needs to be clarified. So, this study aimed to investigate the effect of LXRs activation on testicular changes in rats exposed to electromagnetic radiation. Methods: Thirty adult male Wistar rats were subdivided into control group, experimental group: exposed to cell phone electromagnetic radiation, and treated group: as experimental group and received LXRs agonist T0901317. Serum testosterone (T), Follicle-Stimulating Hormone (FSH), and Luteinizing Hormone (LH), testicular tissue malondialdehyde (MDA) and superoxide dismutase (SOD), and gene expressions of inflammatory and apoptotic markers were assessed. Seminal analysis and testicular histological examination were performed. Results: LXRs activation in rats exposed to cell phone radiation improved the decreased testicular and gonadotrophic hormones, semen quality parameters, tissue SOD, and anti-apoptotic gene expression as well as the elevated tissue MDA, apoptotic and inflammatory gene expressions, and the testicular histological changes induced by exposure to electromagnetic radiation. Conclusion: Exposure to cell phone radiation leads to harmful effects on spermatogenesis and LXRs activation could improve these effects via antioxidant, anti-inflammatory properties in addition to its endocrinal action.

ApoA5 lowers triglyceride levels via suppression of ANGPTL3/8-mediated LPL inhibition

AbstractTriglyceride (TG) molecules represent the major storage form of fatty acids, and TG metabolism is essential to human health. However, the mechanistic details surrounding TG metabolism are complex and incompletely elucidated. Although it is known that angiopoietin-like protein 8 (ANGPTL8) increases TGs through an ANGPTL3/8 complex that inhibits LPL, the mechanism governing ApoA5, which lowers TGs, has remained elusive. Current hypotheses for how ApoA5 acts include direct stimulation of LPL, facilitation of TG-containing particle uptake, and regulation of hepatic TG secretion. Using immunoprecipitation-MS and Western blotting, biolayer interferometry, functional LPL enzymatic assays, and kinetic analyses of LPL activity, we show that ApoA5 associates with ANGPTL3/8 in human serum and most likely decreases TG by suppressing ANGPTL3/8-mediated LPL inhibition. We also demonstrate that ApoA5 has no direct effect on LPL, nor does it suppress the LPL-inhibitory activities of ANGPTL3, ANGPTL4, or ANGPTL4/8. Importantly, ApoA5 suppression of ANGPTL3/8-mediated LPL inhibition occurred at a molar ratio consistent with the circulating concentrations of ApoA5 and ANGPTL3/8. Because liver X receptor (LXR) agonists decrease ApoA5 expression and cause hypertriglyceridemia, we investigated the effect of the prototypical LXR agonist T0901317 on human primary hepatocytes. We observed that T0901317 modestly stimulated hepatocyte ApoA5 release, but markedly stimulated ANGPTL3/8 secretion. Interestingly, the addition of insulin to T0901317 attenuated ApoA5 secretion, but further increased ANGPTL3/8 secretion. Together, these results reveal a novel intersection of ApoA5 and ANGPTL3/8 in the regulation of TG metabolism and provide a possible explanation for LXR agonist-induced hypertriglyceridemia.

Efferocytosis potentiates the expression of arachidonate 15-lipoxygenase (ALOX15) in alternatively activated human macrophages through LXR activation

Macrophages acquire anti-inflammatory and proresolving functions to facilitate resolution of inflammation and promote tissue repair. While alternatively activated macrophages (AAMs), also referred to as M2 macrophages, polarized by type 2 (Th2) cytokines IL-4 or IL-13 contribute to the suppression of inflammatory responses and play a pivotal role in wound healing, contemporaneous exposure to apoptotic cells (ACs) potentiates the expression of anti-inflammatory and tissue repair genes. Given that liver X receptors (LXRs), which coordinate sterol metabolism and immune cell function, play an essential role in the clearance of ACs, we investigated whether LXR activation following engulfment of ACs selectively potentiates the expression of Th2 cytokine-dependent genes in primary human AAMs. We show that AC uptake simultaneously upregulates LXR-dependent, but suppresses SREBP-2-dependent gene expression in macrophages, which are both prevented by inhibiting Niemann–Pick C1 (NPC1)-mediated sterol transport from lysosomes. Concurrently, macrophages accumulate sterol biosynthetic intermediates desmosterol, lathosterol, lanosterol, and dihydrolanosterol but not cholesterol-derived oxysterols. Using global transcriptome analysis, we identify anti-inflammatory and proresolving genes including interleukin-1 receptor antagonist (IL1RN) and arachidonate 15-lipoxygenase (ALOX15) whose expression are selectively potentiated in macrophages upon concomitant exposure to ACs or LXR agonist T0901317 (T09) and Th2 cytokines. We show priming macrophages via LXR activation enhances the cellular capacity to synthesize inflammation-suppressing specialized proresolving mediator (SPM) precursors 15-HETE and 17-HDHA as well as resolvin D5. Silencing LXRα and LXRβ in macrophages attenuates the potentiation of ALOX15 expression by concomitant stimulation of ACs or T09 and IL-13. Collectively, we identify a previously unrecognized mechanism of regulation whereby LXR integrates AC uptake to selectively shape Th2-dependent gene expression in AAMs.

Liver X receptor \u03b2 is required for the survival of single-positive thymocytes by regulating IL-7R\u03b1 expression

Liver X receptors (LXRs) are known as key transcription factors in lipid metabolism and have been reported to play an important role in T-cell proliferation. However, whether LXRs play a role in thymocyte development remains largely unknown. Here, we demonstrated that LXRβ deficiency caused a reduction in single-positive (SP) thymocytes, whereas the transitions from the double-negative to SP stage were normal. Meanwhile, LXRβ-null SP thymocytes exhibited increased apoptosis and impairment of the IL-7Rα-Bcl2 axis. In addition, the LXR agonist T0901317 promoted the survival of SP thymocytes with enhanced IL-7Rα expression in wild-type mice but not in LXRβ-deficient mice. Mechanistically, LXRβ positively regulated the expression of IL-7Rα via direct binding to the Il7r allele in SP thymocytes, and forced expression of IL-7Rα or Bcl2 restored the survival of LXRβ-defective SP thymocytes. Thus, our results indicate that LXRβ functions as an important transcription factor upstream of IL-7Rα to promote the survival of SP thymocytes.

Reduced Sirtuin1 signalling exacerbates diabetic mice hindlimb ischaemia injury and inhibits the protective effect of a liver X receptor agonist

Diabetes mellitus causes endothelial dysfunction, which further exacerbates peripheral arterial disease (PAD). Improving endothelial function via reducing endothelial oxidative stress (OS) may be a promising therapy for diabetic PAD. Activation of liver X receptor (LXR) inhibits excessive OS and provides protective effects on endothelial cells in diabetic individuals. Therefore, we investigated the effects of LXR agonist treatment on diabetic PAD with a focus on modulating endothelial OS. We used a streptozotocin‐induced diabetes mouse model combined with a hindlimb ischaemia (HLI) injury to mimic diabetic PAD, which was followed by LXR agonist treatment. In our study, the LXR agonist T0901317 protected against HLI injury in diabetic mice by attenuating endothelial OS and stimulating angiogenesis. However, a deficiency in endothelial Sirtuin1 (SIRT1) largely inhibited the therapeutic effects of T0901317. Furthermore, we found that the underlying therapeutic mechanisms of T0901317 were related to SIRT1 and non‐SIRT1 signalling, and the isoform LXRβ was involved in LXR agonist‐elicited SIRT1 regulation. In conclusion, LXR agonist treatment protected against HLI injury in diabetic mice via mitigating endothelial OS and stimulating cellular viability and angiogenesis by LXRβ, which elicited both SIRT1‐mediated and non‐SIRT1‐mediated signalling pathways. Therefore, LXR agonist treatment may be a promising therapeutic strategy for diabetic PAD.

Differential Role of Liver X Receptor (LXR) α and LXRβ in the Regulation of UDP-Glucuronosyltransferase 1A1 in Humanized UGT1 Mice.

Liver X receptors (LXRs), LXRα and LXRβ, are nuclear receptors that regulate the metabolism of cholesterol and bile acids and are activated by oxysterols. Humanized UGT1 (hUGT1) mice express the 9-human UGT1A genes associated with the UGT1 locus in a Ugt1-null background. The expression of UGT1A1 is developmentally delayed in the liver and intestines, resulting in the accumulation of serum bilirubin during the neonatal period. Induction of UGT1A1 in newborn hUGT1 mice leads to rapid reduction in total serum bilirubin (TSB) levels, a phenotype measurement that allows for an accurate prediction on UGT1A1 expression. When neonatal hUGT1 mice were treated by oral gavage with the LXR agonist T0901317, TSB levels were dramatically reduced. To determine the LXR contribution to the induction of UGT1A1 and the lowering of TSB levels, experiments were conducted in neonatal hUGT1/Lxrα -/- , hUGT1/Lxrβ -/- , and hUGT1/Lxrαβ -/- mice treated with T0901317. Induction of liver UGT1A1 was dependent upon LXRα, with the induction pattern paralleling induction of LXRα-specific stearoyl CoA desaturase 1. However, the actions of T0901317 were also shown to display a lack of specificity for LXR, with the induction of liver UGT1A1 in hUGT1/Lxrαβ -/- mice, a result associated with activation of both pregnane X receptor and constitutive androstane receptor. However, the LXR agonist GW3965 was highly selective toward LXRα, showing no impact on lowering TSB values or inducing UGT1A1 in hUGT1/Lxrα -/- mice. An LXR-specific enhancer site on the UGT1A1 gene was identified, along with convincing evidence that LXRα is crucial in maintaining constitutive expression of UGT1A1 in adult hUGT1 mice. SIGNIFICANCE STATEMENT: It has been established that activation of LXRα, and not LXRβ, is responsible for the induction of liver UGT1A1 and metabolism of serum bilirubin in neonatal hUGT1 mice. Although induction of the human UGT1A1 gene is initiated at a newly characterized LXR enhancer site, allelic deletion of the Lxrα gene drastically reduces the constitutive expression of liver UGT1A1 in adult hUGT1 mice. Combined, these findings indicate that LXRα is critical for the developmental expression of UGT1A1.

The effects of LXR agonist T0901317 and LXR antagonist GSK2033 on morphogenesis and lipid properties in full thickness skin models

Full thickness models (FTMs) are 3D-cultured human skin models that mimic many aspects of native human skin (NHS). However, their stratum corneum (SC) lipid composition differs from NHS causing a reduced skin barrier. The most pronounced differences in lipid composition are a reduction in lipid chain length and increased monounsaturated lipids. The liver-X-receptor (LXR) activates the monounsaturated lipid synthesis via stearoyl-CoA desaturase-1 (SCD-1). Therefore, the aim was to improve the SC lipid synthesis of FTMs by LXR deactivation. This was achieved by supplementing culture medium with LXR antagonist GSK2033. LXR agonist T0901317 was added for comparison. Subsequently, epidermal morphogenesis, lipid composition, lipid organization and the barrier functionality of these FTMs were assessed. We demonstrate that LXR deactivation resulted in a lipid composition with increased overall chain lengths and reduced levels of monounsaturation, whereas LXR activation increased the amount of monounsaturated lipids and led to a reduction in the overall chain length. However, these changes did not affect the barrier functionality. In conclusion, LXR deactivation led to the development of FTMs with improved lipid properties, which mimic the lipid composition of NHS more closely. These novel findings may contribute to design interventions to normalize SC lipid composition of atopic dermatitis patients.

PNPLA3 I148M Variant Impairs Liver X Receptor Signaling and Cholesterol Homeostasis in Human Hepatic Stellate Cells.

The patatin‐like phospholipase domain‐containing protein 3 (PNPLA3) I148M variant predisposes to hepatic steatosis and progression to advanced liver injury with development of fibrosis, cirrhosis, and cancer. Hepatic stellate cells (HSCs) drive the wound healing response to chronic injury, and lack of liver X receptor (LXR) signaling exacerbates liver fibrogenesis by impairing HSC cholesterol homeostasis. However, the contribution of the I148M variant to this process is still unknown. We analyzed LXR expression and transcriptional activity in primary human HSCs and overexpressing LX‐2 cells according to PNPLA3 genotype (wild type [WT] versus I148M). Here we demonstrate that LXRα protein increased whereas LXR target gene expression decreased during in vitro activation of primary human HSCs. Notably, LXRα levels and signaling were reduced in primary I148M HSCs compared to WT, as displayed by decreased expression of LXR target genes. Moreover, reduced expression of cholesterol efflux and enzymes generating oxysterols was associated with higher total and free cholesterol accumulation whereas endogenous cholesterol synthesis and uptake were diminished in I148M HSCs. Luciferase assays on LXR response element confirmed decreased LXR transcriptional activity in I148M HSCs; in contrast the synthetic LXR agonist T0901317 replenished LXR functionality, supported by adenosine triphosphate‐binding cassette subfamily A member 1 (ABCA1) induction, and reduced collagen1α1 and chemokine (C‐C motif) ligand 5 expression. Conversely, the peroxisome proliferator‐activated receptor gamma (PPARγ) agonist rosiglitazone had only partial effects on the LXR target gene ABCA1, and neither diminished expression of proinflammatory cytokines nor increased de novo lipogenic genes in I148M HSCs. Conclusion: As a consequence of reduced PPARγ activity, HSCs carrying I148M PNPLA3 show impaired LXR signaling, leading to cholesterol accumulation. The use of a specific LXR agonist shows beneficial effects for diminishing sustained HSC activation and development of liver fibrogenesis.

Abstract 4377: Liver X receptor mediated lipotoxicity represents a treatment option for liver cancer

Abstract Solid tumors evolve significant changes in metabolic pathways during development by virtue of their specific biosynthetic demands, their particular microenvironment and the potential occurrence of toxic metabolites such as reactive oxygen species. However, the development of cancer treatment approaches that are based on the inhibition of biosynthetic routes is impaired due to the high plasticity of metabolic networks, e.g. resulting in activation of compensatory pathways or in an increased exchange of metabolites between cancer cells and the tumor environment. Therefore, such therapies could not be translated into efficient clinical applications so far. Here we show that an enhanced lipogenesis, triggered by a pharmacological activation of the Liver X receptor (LXR), represents a new therapeutic strategy for the treatment of liver carcinoma (HCC). A combination of LXR mediated fatty acid synthesis and concomitant Raf suppression results in oxidative stress, induction of a critical ER stress response and subsequently in apoptosis of different murine and human liver cancer cells. Our mechanistic studies identified Raf as an important regulator of lipid metabolism in liver cancer. We found that Raf-1 directly interacts with Stearoyl-CoA desaturase-1 (Scd1), the central enzyme for the conversion of saturated into mono-unsaturated fatty acids and thereby maintains Scd1 protein stability in HCC cells. Thus, inhibition of Raf by Sorafenib diminished Scd1 protein abundance leading to toxic accumulation of saturated fatty acids and metabolic stress in cancer cells under sustained lipogenesis. Treatment studies in autochthonous liver cancer mouse models and xenograft models of human HCC revealed that a combinatorial therapy, consisting of the LXR agonist T0901317 and Sorafenib is highly potent to suppress liver cancer development and to extend the survival of tumor bearing animals. Such a therapy was efficient against hepatic neoplasia with different metabolic phenotypes and well tolerated by mice, even by animals that already suffer from a fatty liver disease. Taken together, we here propose a pharmacologically induced accumulation of toxic metabolites in cancer cells as a new strategy for efficient metabolic targeting of therapy refractory solid tumors. Citation Format: Ramona Rudalska, Jule Harbig, Marteinn Snaebjoernsson, Lyudmyla Taranets, Florian Heinzmann, Stefan Zwirner, Wei Ciu Hu, Thales Kronenberger, Tae-Won Kang, Antti Poso, Stefan Laufer, Mathias Rosenfeldt, Nisar P. Malek, Bernd Pichler, Nikita Popov, Almut Schulze, Lars Zender, Daniel Dauch. Liver X receptor mediated lipotoxicity represents a treatment option for liver cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4377.

Sirt1 counteracts decrease in membrane phospholipid unsaturation and diastolic dysfunction during saturated fatty acid overload

BackgroundThe fatty acid (FA) composition of membrane phospholipid reflects at least in part dietary fat composition. Saturated FA (SFA) suppress Sirt1 activity, while monounsaturated FA (MUFA) counteract this effect. ObjectiveWe explored a role of Sirt1 in homeostatic control of the fatty acid composition of membrane phospholipid in the presence of SFA overload. Methods and resultsSirt1 deficiency in cardiomyocytes decreased the expression levels of liver X receptor (LXR)-target genes, particularly stearoyl-CoA desaturase-1 (Scd1), a rate-limiting enzyme in the cellular synthesis of MUFA from SFA, increased membrane SFA/MUFA ratio, and worsened left ventricular (LV) diastolic function in mice fed an SFA-rich high fat diet. In cultured cardiomyocytes, Sirt1 knockdown (KD) exacerbated the palmitate overload-induced increase in membrane SFA/MUFA ratio, which was associated with decrease in the expression of LXR-target genes, including Scd1. Forced overexpression of Scd1 in palmitate-overloaded Sirt1KD cardiomyocytes lowered the SFA/MUFA ratio. Nicotinamide mononucleotide (NMN) increased Sirt1 activity and Scd1 expression, thereby lowering membrane SFA/MUFA ratio in palmitate-overloaded cardiomyocytes. These effects of NMN were not observed for Scd1KD cardiomyocytes. LXRα/βKD exacerbated palmitate overload-induced increase in membrane SFA/MUFA ratio, while LXR agonist T0901317 alleviated it. NMN failed to rescue Scd1 protein expression and membrane SFA/MUFA ratio in palmitate-overloaded LXRα/βKD cardiomyocytes. The administration of NMN or T0901317 showed a dramatic reversal in membrane SFA/MUFA ratio and LV diastolic function in SFA-rich HFD-fed mice. ConclusionCardiac Sirt1 counteracted SFA overload-induced decrease in membrane phospholipid unsaturation and diastolic dysfunction via regulating LXR-mediated transcription of the Scd1 gene.